Specific human secretory phospholipas A2 (sPLA2)and sPLA2-X were found to neutralize HIV-1 through degradation of the viral membrane. Catalytic function was required for anti-viral activity, and target cells of infection were unaffected. sPLA2-X potently reduced gene transfer of HIV-1 Env-pseudotyped lentiviral vectors and inhibited both CCR5- and CXCR4-tropic HIV-1 virus replication in human CD4+ T cells. Virions resistant to damage by antibody and complement were sensitive to lysis by sPLA2-X, suggesting a novel mechanism of antiviral surveillance independent of the acquired immune system.